Immunogenicity of a Plasmodium vivax vaccine based on the duffy binding protein formulated using adjuvants compatible for use in humans.

The invasion of reticulocytes by Plasmodium vivax merozoites is dependent on the interaction of the Plasmodium vivax Duffy Binding Protein (PvDBP) with the Duffy antigen receptor for chemokines (DARC). The N-terminal cysteine-rich region II of PvDBP (PvDBPII), which binds DARC, is a leading P. vivax malaria vaccine candidate. Here, we have evaluated the immunogenicity of recombinant PvDBPII formulated with the adjuvants Matrix-M and GLA-SE in mice. Analysis of the antibody responses revealed comparable ELISA recognition titres as well as similar recognition of native PvDBP in P. vivax schizonts by immunofluorescence assay. Moreover, antibodies elicited by the two adjuvant formulations had similar functional properties such as avidity, isotype profile and inhibition of PvDBPII-DARC binding. Furthermore, the anti-PvDBPII antibodies were able to block the interaction of DARC with the homologous PvDBPII SalI allele as well as the heterologous PvDBPII PvW1 allele from a Thai clinical isolate that is used for controlled human malaria infections (CHMI). The cross-reactivity of these antibodies with PvW1 suggest that immunization with the PvDBPII SalI strain should neutralize reticulocyte invasion by the challenge P. vivax strain PvW1.

T cell migration and effector function differences in familial adenomatous polyposis patients with APC gene mutations.

Familial adenomatous polyposis (FAP) is an inherited disease characterized by the development of large number of colorectal adenomas with high risk of evolving into colorectal tumors. Mutations of the Adenomatous polyposis coli (APC) gene is often at the origin of this disease, as well as of a high percentage of spontaneous colorectal tumors. APC is therefore considered a tumor suppressor gene. While the role of APC in intestinal epithelium homeostasis is well characterized, its importance in immune responses remains ill defined. Our recent work indicates that the APC protein is involved in various phases of both CD4 and CD8 T cells responses. This prompted us to investigate an array of immune cell features in FAP subjects carrying APC mutations. A group of 12 FAP subjects and age and sex-matched healthy controls were studied. We characterized the immune cell repertoire in peripheral blood and the capacity of immune cells to respond ex vivo to different stimuli either in whole blood or in purified T cells. A variety of experimental approaches were used, including, pultiparamater flow cytometry, NanosString gene expression profiling, Multiplex and regular ELISA, confocal microscopy and computer-based image analyis methods. We found that the percentage of several T and natural killer (NK) cell populations, the expression of several genes induced upon innate or adaptive immune stimulation and the production of several cytokines and chemokines was different. Moreover, the capacity of T cells to migrate in response to chemokine was consistently altered. Finally, immunological synapses between FAP cytotoxic T cells and tumor target cells were more poorly structured. Our findings of this pilot study suggest that mild but multiple immune cell dysfunctions, together with intestinal epithelial dysplasia in FAP subjects, may facilitate the long-term polyposis and colorectal tumor development. Although at an initial discovery phase due to the limited sample size of this rare disease cohort, our findings open new perspectives to consider immune cell abnormalities into polyposis pathology.

Complement-dependent mpox-virus-neutralizing antibodies in infected and vaccinated individuals.

Mpox virus (MPXV) caused a multi-country outbreak in non-endemic areas in 2022. Following historic success of smallpox vaccination with vaccinia virus (VACV)-based vaccines, the third generation modified vaccinia Ankara (MVA)-based vaccine was used as prophylaxis for MPXV, but its effectiveness remains poorly characterized. Here, we applied two assays to quantify neutralizing antibodies (NAbs) in sera from control, MPXV-infected, or MVA-vaccinated individuals. Various levels of MVA NAbs were detected after infection, historic smallpox, or recent MVA vaccination. MPXV was minimally sensitive to neutralization. However, addition of complement enhanced detection of responsive individuals and NAb levels. Anti-MVA and -MPXV NAbs were observed in 94% and 82% of infected individuals, respectively, and 92% and 56% of MVA vaccinees, respectively. NAb titers were higher in individuals born before 1980, highlighting the impact of historic smallpox vaccination on humoral immunity. Altogether, our results indicate that MPXV neutralization is complement dependent and uncover mechanisms underlying vaccine effectiveness.

The tumor suppressor adenomatous polyposis coli regulates T lymphocyte migration.

Adenomatous polyposis coli (APC) is a tumor suppressor whose mutations underlie familial adenomatous polyposis (FAP) and colorectal cancer. Although its role in intestinal epithelial cells is well characterized, APC importance in T cell biology is ill defined. APC regulates cytoskeleton organization, cell polarity, and migration in various cell types. Here, we address whether APC plays a role in T lymphocyte migration. Using a series of cell biology tools, we unveiled that T cells from FAP patients carrying APC mutations display impaired adhesion and motility in constrained environments. We further dissected the cellular mechanisms underpinning these defects in APC-depleted CEM T cell line that recapitulate the phenotype observed in FAP T cells. We found that APC affects T cell motility by modulating integrin-dependent adhesion and cytoskeleton reorganization. Hence, APC mutations in FAP patients not only drive intestinal neoplasms but also impair T cell migration, potentially contributing to inefficient antitumor immunity.

Sustained high expression of multiple APOBEC3 cytidine deaminases in systemic lupus erythematosus.

APOBEC3 (A3) enzymes are best known for their role as antiviral restriction factors and as mutagens in cancer. Although four of them, A3A, A3B, A3F and A3G, are induced by type-1-interferon (IFN-I), their role in inflammatory conditions is unknown. We thus investigated the expression of A3, and particularly A3A and A3B because of their ability to edit cellular DNA, in Systemic Lupus Erythematosus (SLE), a chronic inflammatory disease characterized by high IFN-α serum levels. In a cohort of 57 SLE patients, A3A and A3B, but also A3C and A3G, were upregulated ~ 10 to 15-fold (> 1000-fold for A3B) compared to healthy controls, particularly in patients with flares and elevated serum IFN-α levels. Hydroxychloroquine, corticosteroids and immunosuppressive treatment did not reverse A3 levels. The A3AΔ3B polymorphism, which potentiates A3A, was detected in 14.9% of patients and in 10% of controls, and was associated with higher A3A mRNA expression. A3A and A3B mRNA levels, but not A3C or A3G, were correlated positively with dsDNA breaks and negatively with lymphopenia. Exposure of SLE PBMCs to IFN-α in culture induced massive and sustained A3A levels by 4 h and led to massive cell death. Furthermore, the rs2853669 A > G polymorphism in the telomerase reverse transcriptase (TERT) promoter, which disrupts an Ets-TCF-binding site and influences certain cancers, was highly prevalent in SLE patients, possibly contributing to lymphopenia. Taken together, these findings suggest that high baseline A3A and A3B levels may contribute to cell frailty, lymphopenia and to the generation of neoantigens in SLE patients. Targeting A3 expression could be a strategy to reverse cell death and the generation of neoantigens.

Investigation of viral etiology in potentially malignant disorders and oral squamous cell carcinomas in non-smoking, non-drinking patients.

Head and neck squamous cell carcinomas (HNSCC) are the seventh most frequent cancers. Among HNSCCs, oral squamous cell carcinomas (OSCCs) include several anatomical locations of the oral cavity, but exclude the oropharynx. The known risk factors for OSCCs are mainly alcohol consumption and tobacco use for at least 75-80% of cases. In addition to these risk factors, Human papillomavirus (HPV) types 16 and 18, classified as high-risk (HR) HPV genotypes, are considered as risk factors for oropharyngeal cancers, but their role in the development of OSCC remains unclear. We tested the hypothesis of viral etiology in a series of 68 well-characterized OSCCs and 14 potentially malignant disorders (PMD) in non-smoking, non-drinking (NSND) patients using broad-range, sensitive molecular methodologies. Deep-sequencing of the transcriptome did not reveal any vertebrate virus sequences other than HPV transcripts, detected in only one case. In contrast, HPV DNA was detected in 41.2% (28/68) and 35.7% (5/14) of OSCC and PMD cases, respectively. Importantly, 90.9% (30/33) of these belonged to the Betapapillomavirus genus, but no viral transcripts were detected. Finally, high-throughput sequencing revealed reads corresponding to transcripts of the Trichomonas vaginalis virus (TVV), which were confirmed by RT-PCR in two OSCCs. Our results strongly suggest that Alphapapillomavirus genotypes classified as HR are not involved in the development of OSCCs in NSND patients and that known oncogenic infectious agents are absent in these specific OSCCs. Any possible direct or indirect role of Betapapillomavirus genus members and TVV in OSCCs remains speculative and requires further investigation.

Broad-Range Papillomavirus Transcriptome as a Biomarker of Papillomavirus-Associated Cervical High-Grade Cytology.

Human papillomaviruses (HPVs) are responsible for >99% of cervical cancers. Molecular diagnostic tests based on the detection of viral DNA or RNA have low positive predictive values for the identification of cancer or precancerous lesions. Triage with the Papanicolaou test lacks sensitivity; and even when combined with molecular detection of high-risk HPV, this results in a significant number of unnecessary colposcopies. We have developed a broad-range detection test of HPV transcripts to take a snapshot of the transcriptome of 16 high-risk or putative high-risk HPVs in cervical lesions (HPVs 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, 68, 73, and 82). The purpose of this novel molecular assay, named HPV RNA-Seq, is to detect and type HPV-positive samples and to determine a combination of HPV reads at certain specific viral spliced junctions that can better correlate with high-grade cytology, reflecting the presence of precancerous cells. In a proof-of-concept study conducted on 55 patients, starting from cervical smears, we have shown that HPV RNA-Seq can detect papillomaviruses with performances comparable to a widely used HPV reference molecular diagnostic kit; and a combination of the number of sequencing reads at specific early versus late HPV transcripts can be used as a marker of high-grade cytology, with encouraging diagnostic performances as a triage test.

The rabbit as an orthologous small animal model for APOBEC3A oncogenesis.

APOBEC3 are cytidine deaminases that convert cytidine to uridine residues. APOBEC3A and APOBEC3B enzymes able to target genomic DNA are involved in oncogenesis of a sizeable proportion of human cancers. While the APOBEC3 locus is conserved in mammals, it encodes from 1-7 genes. APOBEC3A is conserved in most mammals, although absent in pigs, cats and throughout Rodentia whereas APOBEC3B is restricted to the Primate order. Here we show that the rabbit APOBEC3 locus encodes two genes of which APOBEC3A enzyme is strictly orthologous to human APOBEC3A. The rabbit enzyme is expressed in the nucleus and the cytoplasm, it can deaminate cytidine, 5-methcytidine residues, nuclear DNA and induce double-strand DNA breaks. The rabbit APOBEC3A enzyme is negatively regulated by the rabbit TRIB3 pseudokinase protein which is guardian of genome integrity, just like its human counterpart. This indicates that the APOBEC3A/TRIB3 pair is conserved over approximately 100 million years. The rabbit APOBEC3A gene is widely expressed in rabbit tissues, unlike human APOBEC3A. These data demonstrate that rabbit could be used as a small animal model for studying APOBEC3 driven oncogenesis.

Self-cytoplasmic DNA upregulates the mutator enzyme APOBEC3A leading to chromosomal DNA damage.

Foreign and self-cytoplasmic DNA are recognized by numerous DNA sensor molecules leading to the production of type I interferons. Such DNA agonists should be degraded otherwise cells would be chronically stressed. Most human APOBEC3 cytidine deaminases can initiate catabolism of cytoplasmic mitochondrial DNA. Using the human myeloid cell line THP-1 with an interferon inducible APOBEC3A gene, we show that cytoplasmic DNA triggers interferon α and ß production through the RNA polymerase III transcription/RIG-I pathway leading to massive upregulation of APOBEC3A. By catalyzing C→U editing in single stranded DNA fragments, the enzyme prevents them from re-annealing so attenuating the danger signal. The price to pay is chromosomal DNA damage in the form of CG→TA mutations and double stranded DNA breaks which, in the context of chronic inflammation, could drive cells down the path toward cancer.

Molecular basis of the attenuated phenotype of human APOBEC3B DNA mutator enzyme.

The human APOBEC3A and APOBEC3B genes (A3A and A3B) encode DNA mutator enzymes that deaminate cytidine and 5-methylcytidine residues in single-stranded DNA (ssDNA). They are important sources of mutations in many cancer genomes which show a preponderance of CG->TA transitions. Although both enzymes can hypermutate chromosomal DNA in an experimental setting, only A3A can induce double strand DNA breaks, even though the catalytic domains of A3B and A3A differ by only 9% at the protein level. Accordingly we sought the molecular basis underlying A3B attenuation through the generation of A3A-A3B chimeras and mutants. It transpires that the N-terminal domain facilitates A3B activity while a handful of substitutions in the catalytic C-terminal domain impacting ssDNA binding serve to attenuate A3B compared to A3A. Interestingly, functional attenuation is also observed for the rhesus monkey rhA3B enzyme compared to rhA3A indicating that this genotoxic dichotomy has been selected for and maintained for some 38 million years. Expression of all human ssDNA cytidine deaminase genes is absent in mature sperm indicating they contribute to somatic mutation and cancer but not human diversity.

The Merkel Cell Polyomavirus: an oncogenic virus infecting humans.

The Merkel cell polyomavirus has been named after a rare type of skin cancer, Merkel cell carcinoma, in which the virus was identified in 2008. It belongs to a family that includes well-known carcinogenic viruses in animals. Five years later, data are now available on Merkel cell polyomavirus structure, genomic organization, protein functions and replication cycle although there is still a need of in vitro and in vivo models. Despite the ubiquitous aspect of the MCPyV infection, the identification of several tumour specific viral markers supports that Merkel cell polyomavirus is indeed an etiologic agent of Merkel cell carcinoma. Moreover, whereas some mechanisms previously described with others polyomavirus have been confirmed, some new interactions between cellular and viral proteins have been identified that pave the way to the understanding of the molecular mechanisms underlying cellular transformation. Finally, in a clinical perspective, Merkel cell polyomavirus research brings potential tools for diagnosis, prognosis evaluation and new therapeutic approaches to manage Merkel cell carcinoma, a severe disease the frequency of which is increasing.

Trichodysplasia spinulosa associated with lupus.

Trichodysplasia spinulosa (TS) is a unique clinical and histological entity described in immunosuppressed patients. The recent discovery of genomic DNA from a new Polyomavirus named trichodysplasia spinulosa-associated Polyomavirus in TS lesions and good clinical response to cidofovir strengthens the hypothesis of a viral etiology for the disease. The authors report a case of TS associated with lupus erythematosus in a 26-year-old woman with no history of transplant, hemopathy, or cyclosporine treatment. The patient developed a progressive worsening eruption composed of confluent papules and spiky filiform excrescences concentrated in the midfacial area. Pathological features were characterized by aberrant distended and abnormally maturated hair follicles with sheets of eosinophilic cells containing large purple granules, and the presence of trichodysplasia spinulosa-associated Polyomavirus DNA was confirmed by polymerase chain reaction. This case is the first description in a nontransplanted lupus patient without underlying hemopathy.

Arsenic trioxide prevents murine sclerodermatous graft-versus-host disease.

Chronic graft-versus-host disease (GVHD) follows allogeneic hematopoietic stem cell transplantation. It results from alloreactive processes induced by minor MHC incompatibilities triggered by activated APCs, such as plasmacytoid dendritic cells (pDCs), and leading to the activation of CD4 T cells. Therefore, we tested whether CD4(+) and pDCs, activated cells that produce high levels of reactive oxygen species, could be killed by arsenic trioxide (As(2)O(3)), a chemotherapeutic drug used in the treatment of acute promyelocytic leukemia. Indeed, As(2)O(3) exerts its cytotoxic effects by inducing a powerful oxidative stress that exceeds the lethal threshold. Sclerodermatous GVHD was induced in BALB/c mice by body irradiation, followed by B10.D2 bone marrow and spleen cell transplantation. Mice were simultaneously treated with daily i.p. injections of As(2)O(3). Transplanted mice displayed severe clinical symptoms, including diarrhea, alopecia, vasculitis, and fibrosis of the skin and visceral organs. The symptoms were dramatically abrogated in mice treated with As(2)O(3). These beneficial effects were mediated through the depletion of glutathione and the overproduction of H(2)O(2) that killed activated CD4(+) T cells and pDCs. The dramatic improvement provided by As(2)O(3) in the model of sclerodermatous GVHD that associates fibrosis with immune activation provides a rationale for the evaluation of As(2)O(3) in the management of patients affected by chronic GVHD.

Anogenital pseudotumoral herpes and HIV infection: a new challenge for diagnosis and treatment.

HIV-infected patients may develop rare anogenital pseudotumoral herpes potentially mimicking epidermoid carcinoma. We assessed treatment in five new cases with a median follow-up of 3.3 years. Recurrence and clinical nucleoside analog resistance were observed in all patients. All drug treatments were only temporarily curative and clinical responses varied between patients and recurrences. Foscavir seemed to be the most appropriate second-line treatment and cidofovir or thalidomide should be considered as alternative treatments.

High levels of antibodies against merkel cell polyomavirus identify a subset of patients with merkel cell carcinoma with better clinical outcome.

PURPOSE: A new human polyomavirus, Merkel cell polyomavirus (MCV), was identified in 2008 in tumor tissue of patients with Merkel cell carcinoma (MCC), a relatively rare human skin cancer. In this study, we investigated patients with MCC and controls for the presence of antibodies against MCV and their association with clinical characteristics. PATIENTS AND METHODS: Antibodies against MCV were investigated by enzyme-linked immunosorbent assay in 68 patients with MCC and 82 controls using VP1 virus-like particles produced in insect cells. RESULTS: Antibodies against MCV were detected in all patients with MCC and in 85% of controls. However, high antibody titers (> 10,000) were rarely observed in controls (7.3%) and they were detected in 64.7% of patients with MCC (P < .001) in contrast to the absence of VP1 expression in tumor samples. In addition, the geometric mean titer of anti-MCV in patients with MCC was around 14 times higher than that observed in MCV-positive controls (P < .001) and was not correlated with tumor viral load. High antibody titers were not found to be associated with any subject or tumor characteristics, but better progression-free survival was observed in patients with high antibody titers (hazard ratio, 4.6; 95% CI, 1.7 to 12.2; P = .002). CONCLUSION: High titers of MCV antibodies in a much higher proportion of patients with MCC than in controls confirmed the association between MCV infection and MCC. The findings also indicated that a better progression-free survival occurred in patients with high MCV antibody titers and suggested that there are at least two distinct etiologic causes of MCC.

Epidermodysplasia verruciformis in human immunodeficiency virus-infected patients: a marker of human papillomavirus-related disorders not affected by antiretroviral therapy.

BACKGROUND: Skin eruptions resembling epidermodysplasia verruciformis (EV) are rarely observed in immunocompromised patients. We focused on the epidemiologic, clinical, virologic, and immunologic features of EV in human immunodeficiency virus (HIV)-positive patients. OBSERVATIONS: We studied 11 HIV-positive patients (6 men and 5 women) with clinical and histological features of EV observed at our department. The median age at HIV diagnosis was 27 years. At the onset of eruption, the median age was 40 years and the median CD4 T-cell count was 170/µL. Clinical presentation included flat warts (n = 11), pityriasis versicolor-like macules (n = 5), and lichenoid papules (n = 3) on sun-exposed skin. Detection and typing of cutaneous human papillomavirus (HPV) were carried out in 8 cases and always revealed ß-HPV infection, including oncogenic HPV-5 or 8 (n = 6). Mucosal HPV-related diseases were present in 7 cases. Histories of skin cancer and lymphoproliferative disorder were recorded in 3 and 4 patients, respectively, including 2 fatal cases. Skin eruption was never improved by highly active antiretroviral therapy (HAART). In 2 cases, EV was associated with an immune reconstitution syndrome. The present series is the largest with a complete characterization. A review of similar cases was carried out. CONCLUSION: Despite effective HAART, HIV-infected patients with EV require a prolonged and careful follow-up to detect mucosal HPV-related diseases, lymphoproliferative disorders, and skin cancers.

Lipschütz genital ulceration associated with mumps.

Lipschütz ulcers are characterised by a first flare of non-sexually related acute genital ulcers (AGU) occurring in adolescent girls. Epstein-Barr primary infection is the most frequently reported aetiology but other infectious agents are probably implicated. We report the first case of mumps associated with an AGU in a 21-year-old girl. She presented a bilateral parotitis with genital ulcers, and serology confirmed she had mumps. As in our case, most Lipschütz ulcers heal spontaneously within a couple of weeks and the diagnosis should be reconsidered in case of recurrence.

Distinct merkel cell polyomavirus molecular features in tumour and non tumour specimens from patients with merkel cell carcinoma.

Merkel Cell Polyomavirus (MCPyV) is associated with Merkel Cell carcinoma (MCC), a rare, aggressive skin cancer with neuroendocrine features. The causal role of MCPyV is highly suggested by monoclonal integration of its genome and expression of the viral large T (LT) antigen in MCC cells. We investigated and characterized MCPyV molecular features in MCC, respiratory, urine and blood samples from 33 patients by quantitative PCR, sequencing and detection of integrated viral DNA. We examined associations between either MCPyV viral load in primary MCC or MCPyV DNAemia and survival. Results were interpreted with respect to the viral molecular signature in each compartment. Patients with MCC containing more than 1 viral genome copy per cell had a longer period in complete remission than patients with less than 1 copy per cell (34 vs 10 months, P = 0.037). Peripheral blood mononuclear cells (PBMC) contained MCPyV more frequently in patients sampled with disease than in patients in complete remission (60% vs 11%, P = 0.00083). Moreover, the detection of MCPyV in at least one PBMC sample during follow-up was associated with a shorter overall survival (P = 0.003). Sequencing of viral DNA from MCC and non MCC samples characterized common single nucleotide polymorphisms defining 8 patient specific strains. However, specific molecular signatures truncating MCPyV LT were observed in 8/12 MCC cases but not in respiratory and urinary samples from 15 patients. New integration sites were identified in 4 MCC cases. Finally, mutated-integrated forms of MCPyV were detected in PBMC of two patients with disseminated MCC disease, indicating circulation of metastatic cells. We conclude that MCPyV molecular features in primary MCC tumour and PBMC may help to predict the course of the disease.